Deciphering the mechanisms associated with high-risk AMD genotypes for ARMS2/HTRA1 andComplement Factor H

破译与 ARMS2/HTRA1 和补体因子 H 的高风险 AMD 基因型相关的机制

• 批准号: 10707245
• 负责人: Deborah Ann Ferrington
• 金额: $ 38.21万
• 依托单位: DOHENY EYE INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-30 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10707245
• 关键词: Affect; Aftercare; Age related macular degeneration; Applications Grants; Bioinformatics; Biological; Biological Assay; Biological Models; Biological Process; Biology; Cell Line; Cells; Clinical; Clinical Research; Complement Factor H; Coupled; Defect; Elderly; Exhibits; Exposure to; Foundations; Functional disorder; Genes; Genetic; Genetic Risk; Genotype; Goals; Growth Factor; High temperature of physical object; Investigation; Knowledge; Laboratories; Metabolism; Mitochondria; Molecular; Molecular Profiling; New York; Odds Ratio; Oxidative Stress; Parents; Pathogenesis; Pathologic; Pathway interactions; Patient Care; Patient-Focused Outcomes; Patients; Physiological; Population; Predisposition; Proteins; Proteome; Protocols documentation; Risk; Single Nucleotide Polymorphism; Stress; Structure of retinal pigment epithelium; Testing; Thick; Treatment Factor; Vascular Endothelial Growth Factors; biological adaptation to stress; geographic atrophy; high risk; improved; induced pluripotent stem cell; insight; macula; metabolome; neovascularization; novel; patient population; personalized medicine; prototype; response; risk variant; stem cells; targeted treatment; therapy design; therapy development

Project Summary This U01 grant application is submitted in response to RFA EY-21-002 “Age-related Macular Degeneration (AMD) Integrative Biology Initiative”, with the goal of determining if patient-derived induced pluripotent stem cell (iPSC)-retinal pigment epithelium (RPE) can be used to discover the underlying pathophysiology of AMD. Studies from our laboratory as well as those of others strongly support the use of iPSC-RPE to understand how specific single-nucleotide polymorphisms (SNP) associated with high risk for advanced AMD influences multiple biological processes in the RPE. Our studies will focus on two SNPs with the highest odds ratio for developing advanced AMD- Age-Related Maculopathy Susceptibility 2/High Temperature Requirement A1 (ARMS2/HTRA1) and Complement Factor H (CFH). We will use the iPSC developed by the NEI and the New York Stem Cell Foundation (NYSCF) and differentiate the parent cells and their reciprocal isogenic lines into RPE using the protocol established in the Bharti laboratory. The parent/isogenic lines provide an excellent model system for deciphering the molecular factors that determine how the presence of SNPs in one or both genes affect the response to stress. We will test the hypothesis that the presence of homozygous risk alleles for either ARMS2/HTRA1 or CFH, independent and in combination, have a negative impact on the RPE stress response. Furthermore, each genetic risk profile will have a unique stress response. Our proposed studies will integrate classical reductionist investigations, coupled with discovery-driven analyses, to test specific hypotheses. Aim 1 will focus on differentiating and characterizing iPSC-RPE cell lines from 10 AMD patients and their isogenic lines. Aim 2 will test the hypothesis that the presence of AMD high-risk SNPs alter the cellular response to stress by performing large-scale quantification of proteins and metabolites, coupled with mt functional assays. Parent cells and their isogenic lines will be compared to determine the impact on metabolism and the stress response in iPSC-RPE (i) with the CFH high-risk SNP and (ii) for high risk ARMS2/HTRA1, alone or in combination with high risk CFH. These studies will address how the presence of specific risk variants affect RPE exposed to a physiologically relevant stress, an important gap in knowledge of AMD pathogenesis. Knowledge about the molecular details has the potential to lead to development of therapies targeting the primary defect in a genetically defined population of AMD patients, which in turn, could lead to a “personalized medicine” approach for treatment of AMD.

项目摘要 此U 01资助申请是为了响应RFA EY-21-002“视网膜相关黄斑变性”而提交的 (AMD)整合生物学倡议”，目标是确定患者来源的诱导多能干细胞是否 因此，可使用iPSC-视网膜色素上皮（RPE）来发现AMD的潜在病理生理学。 我们实验室的研究以及其他人的研究强烈支持使用iPSC-RPE来了解如何 与晚期AMD高风险相关的特定单核苷酸多态性（SNP）影响多种 RPE中的生物学过程。我们的研究将集中在两个SNPs与发展的优势比最高 晚期AMD-AMD相关黄斑病变易感性2/高温要求A1（ARMS 2/HTRA 1） 和补体因子H（CFH）。我们将使用NEI和纽约干细胞公司开发的iPSC 基础（NYSCF），并使用该方法将亲本细胞和它们的相互等基因系分化成RPE。 在Bharti实验室建立的协议。亲本/等基因系提供了一个极好的模型系统， 破译决定一个或两个基因中SNP的存在如何影响基因表达的分子因素， 对压力的反应。我们将检验以下假设： ARMS 2/HTRA 1或CFH单独或组合对RPE应激反应具有负面影响。 此外，每种遗传风险特征都有独特的压力反应。我们建议的研究将结合 经典的还原主义调查，加上发现驱动的分析，以测试特定的假设。要求1 将专注于区分和表征来自10名AMD患者的iPSC-RPE细胞系及其等基因系。 目的2将检验AMD高危SNP的存在通过以下方式改变细胞对应激的反应的假设： 进行蛋白质和代谢物的大规模定量，并结合MT功能测定。亲本细胞 将比较它们的等基因系，以确定对代谢的影响和 iPSC-RPE（i）具有CFH高风险SNP和（ii）对于高风险ARMS 2/HTRA 1，单独或与高风险SNP组合， 风险CFH。这些研究将解决特定风险变体的存在如何影响暴露于特定环境中的RPE。 生理相关的压力，AMD发病机制的知识的重要差距。的知识 分子细节有可能导致针对原发性缺陷的治疗方法的发展， 基因定义的AMD患者群体，这反过来可能导致“个性化医疗”方法 用于治疗AMD。